Title: Involvement of dopaminergic and cholinergic systems in social isolation-induced deficits in social affiliation and conditional fear memory in mice Okada R, Fujiwara H, Mizuki D, Araki R, Yabe T, Matsumoto K Ref: Neuroscience, 299:134, 2015 : PubMed
Post-weaning social isolation rearing (SI) in rodents elicits various behavioral abnormalities including attention deficit hyperactivity disorder-like behaviors. In order to obtain a better understanding of SI-induced behavioral abnormalities, we herein investigated the effects of SI on social affiliation and conditioned fear memory as well as the neuronal mechanism(s) underlying these effects. Four-week-old male mice were group-housed (GH) or socially isolated for 2-4weeks before the experiments. The social affiliation test and fear memory conditioning were conducted at the age of 6 and 7weeks, respectively. SI mice were systemically administered saline or test drugs 30min before the social affiliation test and fear memory conditioning. Contextual and auditory fear memories were elucidated 1 and 4days after fear conditioning. Social affiliation and contextual and auditory fear memories were weaker in SI mice than in GH mice. Methylphenidate (MPH), an inhibitor for dopamine transporters, ameliorated the SI-induced social affiliation deficit and the effect was attenuated by SCH23390, a D1 receptor antagonist, but not by sulpiride, a D2 receptor antagonist. On the other hand, tacrine, an acetylcholinesterase inhibitor, had no effect on this deficit. In contrast, tacrine improved SI-induced deficits in fear memories in a manner that was reversed by the muscarinic receptor antagonist scopolamine, while MPH had no effect on memory deficits. Neurochemical studies revealed that SI down-regulated the expression levels of the phosphorylated forms of neuro-signaling proteins, calmodulin-dependent kinase II (p-CaMKII), and cyclic AMP-responsive element binding protein (p-CREB), as well as early growth response protein-1 (Egr-1) in the hippocampus. The administration of MPH or tacrine before fear conditioning had no effect on the levels of the phosphorylated forms of the neuro-signaling proteins elucidated following completion of the auditory fear memory test; however, when analyzed 30min after the administration of the test drugs, tacrine significantly attenuated the SI-induced decrease in p-CaMKII, p-CREB, and Egr-1 in a manner reversible by scopolamine. Our results suggest that SI-induced deficits in social affiliation and conditioned fear memory were mediated by functional alterations to central dopaminergic and cholinergic systems, respectively.
        
Title: Butea superba-Induced Amelioration of Cognitive and Emotional Deficits in Olfactory Bulbectomized Mice and Putative Mechanisms Underlying Its Actions Mizuki D, Qi Z, Tanaka K, Fujiwara H, Ishikawa T, Higuchi Y, Matsumoto K Ref: J Pharmacol Sci, 124:457, 2014 : PubMed
This study investigated the effects of alcoholic extract of Butea superba (BS) on cognitive deficits and depression-related behavior using olfactory bulbectomized (OBX) mice and the underlying molecular mechanisms of its actions. OBX mice were treated daily with BS (100 and 300 mg/kg, p.o.) or reference drugs, tacrine (2.5 mg/kg, i.p.) and imipramine (10 mg/kg, i.p.) from day 3 after OBX. OBX impaired non-spatial and spatial cognitive performances, which were elucidated by the novel object recognition test and modified Y maze test, respectively. These deficits were attenuated by tacrine and BS but not imipramine. OBX animals exhibited depression-like behavior in the tail suspension test in a manner reversible by imipramine and BS but not tacrine. OBX down-regulated phosphorylation of synaptic plasticity-related signaling proteins: NMDA receptor, AMPA receptor, calmodulin-dependent kinase II, and cyclic AMP-responsive element-binding protein. OBX also reduced choline acetyltransferase in the hippocampus. BS and tacrine reversed these neurochemical alterations. Moreover, BS inhibited ex vivo activity of acetylcholinesterase in the brain. These results indicate that BS ameliorates not only cognition dysfunction via normalizing synaptic plasticity-related signaling and facilitating central cholinergic systems but also depression-like behavior via a mechanism differing from that implicated in BS amelioration of cognitive function in OBX animals.
In our previous study, the daily administration of chotosan (CTS), a Kampo formula consisting of Uncaria and other 10 different crude drugs, ameliorated cognitive deficits in several animal models of dementia including type 2 diabetic db/db mice in a similar manner to tacrine, an acetylcholinesterase inhibitor. The present study investigated the metabonomics of CTS in db/db mice, a type 2 diabetes model, and m/m mice, a non-diabetes control strain, to identify the exogenous and endogenous chemicals susceptible to the administration of CTS using high performance liquid chromatography equipped with an orbitrap hybrid Fourier transform mass spectrometer. The results obtained revealed that the systemic administration of CTS for 20 days led to the distribution of Uncalia plant-derived alkaloids such as rhynchophylline, hirsuteine, and corynoxeine in the plasma and brains of db/db and m/m mice and induced alterations in four major metabolic pathways; i.e., (1) purine, (2) tryptophan, (3) cysteine and methionine, (4) glycerophospholipids in db/db mice. Moreover, glycerophosphocholine (GPC) levels in the plasma and brain were significantly higher in CTS-treated db/db mice than in vehicle-treated control animals. The results of the in vitro experiment using organotypic hippocampal slice cultures demonstrated that GPC (10-30muM), as well as tacrine, protected hippocampal cells from N-methyl-d-aspartate-induced excitotoxicity in a manner that was reversible with the muscarinic receptor antagonist scopolamine, whereas GPC had no effect on the activity of acetylcholinesterase in vitro. Our results demonstrated that some CTS constituents with neuropharmacological activity were distributed in the plasma and brain tissue following the systemic administration of CTS and may subsequently have affected some metabolic pathways including glycerophospholipid metabolism and cognitive function in db/db mice. Moreover, the present metabonomic analysis suggested that GPC is a putative endogenous chemical that may be involved in the tacrine-like actions of CTS in the present diabetic animal model.
This study investigated the effects of alcoholic extract of Bacopa monnieri (L.) Wettst. (BM) on cognitive deficits using olfactory bulbectomized (OBX) mice and the underlying molecular mechanisms of its action. OBX mice were treated daily with BM (50 mg/kg, p.o.) or a reference drug, tacrine (2.5 mg/kg, i.p.), 1 week before and continuously 3 days after OBX. Cognitive performance of the animals was analyzed by the novel object recognition test, modified Y maze test, and fear conditioning test. Brain tissues of OBX animals were used for neurochemical and immunohistochemical studies. OBX impaired non-spatial short-term memory, spatial working memory, and long-term fair memory. BM administration ameliorated these memory disturbances. The effect of BM on short-term memory deficits was abolished by a muscarinic receptor antagonist, scopolamine. OBX downregulated phosphorylation of synaptic plasticity-related signaling proteins: NR1 subunit of N-methyl-D-aspartate receptor, glutamate receptor 1 (GluR1), and calmodulin-dependent kinase II but not cyclic AMP-responsive element binding protein (CREB), and reduced brain-derived neurotrophic factor (BDNF) mRNA in the hippocampus. OBX also reduced choline acetyltransferase in the hippocampus and cholinergic neurons in the medial septum, and enlarged the size of lateral ventricle. BM administration reversed these OBX-induced neurochemical and histological alterations, except the decrease of GluR1 phosphorylation, and enhanced CREB phosphorylation. Moreover, BM treatment inhibited ex vivo activity of acetylcholinesterase in the brain. These results indicate that BM treatment ameliorates OBX-induced cognition dysfunction via a mechanism involving enhancement of synaptic plasticity-related signaling and BDNF transcription and protection of cholinergic systems from OBX-induced neuronal damage.
        
Title: Comprehensive microarray-based analysis for stage-specific larval camouflage pattern-associated genes in the swallowtail butterfly, Papilio xuthus Futahashi R, Shirataki H, Narita T, Mita K, Fujiwara H Ref: BMC Biol, 10:46, 2012 : PubMed
BACKGROUND: Body coloration is an ecologically important trait that is often involved in prey-predator interactions through mimicry and crypsis. Although this subject has attracted the interest of biologists and the general public, our scientific knowledge on the subject remains fragmentary. In the caterpillar of the swallowtail butterfly Papilio xuthus, spectacular changes in the color pattern are observed; the insect mimics bird droppings (mimetic pattern) as a young larva, and switches to a green camouflage coloration (cryptic pattern) in the final instar. Despite the wide variety and significance of larval color patterns, few studies have been conducted at a molecular level compared with the number of studies on adult butterfly wing patterns. RESULTS: To obtain a catalog of genes involved in larval mimetic and cryptic pattern formation, we constructed expressed sequence tag (EST) libraries of larval epidermis for P. xuthus, and P. polytes that contained 20,736 and 5,376 clones, respectively, representing one of the largest collections available in butterflies. A comparison with silkworm epidermal EST information revealed the high expression of putative blue and yellow pigment-binding proteins in Papilio species. We also designed a microarray from the EST dataset information, analyzed more than five stages each for six markings, and confirmed spatial expression patterns by whole-mount in situ hybridization. Hence, we succeeded in elucidating many novel marking-specific genes for mimetic and cryptic pattern formation, including pigment-binding protein genes, the melanin-associated gene yellow-h3, the ecdysteroid synthesis enzyme gene 3-dehydroecdysone 3b-reductase, and Papilio-specific genes. We also found many cuticular protein genes with marking specificity that may be associated with the unique surface nanostructure of the markings. Furthermore, we identified two transcription factors, spalt and ecdysteroid signal-related E75, as genes expressed in larval eyespot markings. This finding suggests that E75 is a strong candidate mediator of the hormone-dependent coordination of larval pattern formation. CONCLUSIONS: This study is one of the most comprehensive molecular analyses of complicated morphological features, and it will serve as a new resource for studying insect mimetic and cryptic pattern formation in general. The wide variety of marking-associated genes (both regulatory and structural genes) identified by our screening indicates that a similar strategy will be effective for understanding other complex traits.
Tribolium castaneum is a member of the most species-rich eukaryotic order, a powerful model organism for the study of generalized insect development, and an important pest of stored agricultural products. We describe its genome sequence here. This omnivorous beetle has evolved the ability to interact with a diverse chemical environment, as shown by large expansions in odorant and gustatory receptors, as well as P450 and other detoxification enzymes. Development in Tribolium is more representative of other insects than is Drosophila, a fact reflected in gene content and function. For example, Tribolium has retained more ancestral genes involved in cell-cell communication than Drosophila, some being expressed in the growth zone crucial for axial elongation in short-germ development. Systemic RNA interference in T. castaneum functions differently from that in Caenorhabditis elegans, but nevertheless offers similar power for the elucidation of gene function and identification of targets for selective insect control.
Bombyx mori, the domesticated silkworm, is a major insect model for research, and the first lepidopteran for which draft genome sequences became available in 2004. Two independent data sets from whole-genome shotgun sequencing were merged and assembled together with newly obtained fosmid- and BAC-end sequences. The remarkably improved new assembly is presented here. The 8.5-fold sequence coverage of an estimated 432 Mb genome was assembled into scaffolds with an N50 size of approximately 3.7 Mb; the largest scaffold was 14.5 million base pairs. With help of a high-density SNP linkage map, we anchored 87% of the scaffold sequences to all 28 chromosomes. A particular feature was the high repetitive sequence content estimated to be 43.6% and that consisted mainly of transposable elements. We predicted 14,623 gene models based on a GLEAN-based algorithm, a more accurate prediction than the previous gene models for this species. Over three thousand silkworm genes have no homologs in other insect or vertebrate genomes. Some insights into gene evolution and into characteristic biological processes are presented here and in other papers in this issue. The massive silk production correlates with the existence of specific tRNA clusters, and of several sericin genes assembled in a cluster. The silkworm's adaptation to feeding on mulberry leaves, which contain toxic alkaloids, is likely linked to the presence of new-type sucrase genes, apparently acquired from bacteria. The silkworm genome also revealed the cascade of genes involved in the juvenile hormone biosynthesis pathway, and a large number of cuticular protein genes.
During human placentation, the invasion of extravillous trophoblasts (EVTs) into maternal decidual tissues, especially toward maternal spiral arteries, is considered an essential process for subsequent normal fetal development. However, the precise regulatory mechanisms to induce EVT invasion toward arteries and/or to protect EVTs from further invasion have not been well understood. Recently, we found that two cell surface peptidases, dipeptidyl peptidase IV (DPPIV) and carboxypeptidase-M (CP-M,) are differentially expressed on EVTs. DPPIV expression was mainly observed on EVTs that had already ceased invasion. CP-M was detected on migrating EVTs including endovascular trophoblasts in the maternal arteries. The enzymatic inhibition of these peptidases affected the invasive property of choriocarcinoma-derived cell lines, BeWo and JEG3 cells. In addition, a chemokine, RANTES, that is one of the substrates for DPPIV, enhanced invasion of EVTs isolated from primary villous explant culture and its receptor, CCR1, was specifically expressed on migrating EVTs toward maternal arteries. Furthermore, a novel membrane-bound cell surface peptidase, named laeverin, was found to be specifically expressed on EVTs that had almost ceased invasion. These findings suggest that membrane-bound peptidases are important factors regulating EVT invasion during early placentation in humans.
To investigate the mechanisms of ovarian cell differentiation, we raised a new monoclonal antibody, HCL-3, which reacted with human luteal cells. It also reacted with human and porcine hepatocytes. The immunoaffinity-purified HCL-3 antigen from human corpora lutea (CL) was shown to be a 46-kDa protein. The N-terminal 22 amino acids of the 46-kDa protein from porcine liver exhibited high homology (82%) to human microsomal epoxide hydrolase (mEH). The purified HCL-3 antigen from human CL or porcine liver showed EH enzyme activity, confirming that HCL-3 antigen is identical to mEH, which is reported to detoxify the toxic substrates in the liver. In human follicles, mEH was immunohistochemically detected on granulosa and theca interna cells. In the menstrual and pregnant CL, mEH was also expressed on large and small luteal cells. A competitive inhibitor of EH, 1,2-epoxy-3,3,3-trichloropropane, inhibited the conversion of estradiol from testosterone by granulosa cells cultured in vitro, indicating the involvement of mEH in ovarian estrogen production. Because anticonvulsant sodium valproate and its analogues were reported to inhibit EH enzyme activity, these findings provide a new insight into the etiology of endocrine disorders that are frequently observed among epileptic patients taking anticonvulsant drugs.